DNA derived from chromosome band 3 of the cloned Giardia duodenalis line, WB-1B was used to construct a cloned library in E. coli. One of these clones, C3/23, has been identified as the 3' coding region of a G. duodenalis cysteine-rich variable surface protein (CRVSP) gene by homology with other published CRVSPs and also contains 720 bp of the 3' flanking region. The sequence of C3/23, was derived from genomic DNA independently of cDNA, or expression copies of the CRVSP genes. The 3' flanking region is not homologous to the 3' untranslated regions of published CRVSPs which probably reflects its genomic origin. Subclones of C3/23 were used to show that the 3' flanking region was conserved in all strains examined in this study and was repeated many times in the genome. The 3' flanking repeats were located on three chromosome bands and were not always associated with the coding sequence of C3/23 which was represented, although not equally, on all chromosome bands. The highly conserved nature of the 3' flanking region and its multiple representation in the genome emphasize the probable role of this sequence in the localization or regulation of expression of the CRVSPs in G. duodenalis.